The host immune response to periodontal-associated pathogens has been shown to be a key determinant involved in periodontal disease by regulating the production of pro- and anti-inflammatory cytokines that regulate the severity of this disease. An inability to regulate the nature or duration of the host's inflammatory response can often mediate detrimental host effects as observed in chronic inflammatory diseases. Our laboratory has identified the kinase glycogen synthase kinase 3 (GSK3) as a central regulator of the inflammatory process by its ability to differentially regulate the levels of pro- and anti-inflammatory cytokines upon TLR activation. Our specific hypothesis is that the ability of P. gingivalis to interact with TLR2 and TLR4 expressed on immune cells and the subsequent differential regulation of GSK3 activity mediated by these TLRs are major mechanisms responsible for the pathogenesis of this organism by regulating the nature and magnitude of the host's inflammatory response. This hypothesis is based on: 1) stimulation of immune cells from TLR2- or TLR4-deficient mice with P. gingivalis resulted in pronounced differences in the inhibition of GSK3; 2) inhibition of GSK3 in macrophages or dendritic cells stimulated with P. gingivalis resulted in the severe reduction in the levels of pro-inflammatory cytokines, i.e. IL-12, while concurrently augmenting the levels of the anti-inflammatory cytokine IL-10; 3) a direct comparison of TLR2- and TLR4- deficient cells stimulated with P. gingivalis demonstrated that the levels of pro-inflammatory cytokines such as IL-12 were largely mediated by TLR4 whereas the levels of the anti-inflammatory cytokine IL-10 were predominantly mediated by TLR2; 4) analysis of the two major isoforms of GSK3 in innate immune cells revealed that dendritic cells contain only detectable levels of GSKS-p, whereas macrophages contain both isoforms of GSK3, GSK3-a and GSK3-p; 5) in vivo administration of a GSK3 inhibitor resulted in a potent reduction in the levels of IL-1b, IL-6, IL-12, and IFN-y whereas the levels of the anti-inflammatory cytokine IL-10 were potently augmented in response to P. gingivalis; and 6) inhibition of GSK3 in mice resulted in a predominant Th2-assoicated immune response to P. gingivalis. These observations provide a strong rationale and focus concerning the reported differences of TLR2 and TLR4 to mediate pro- and anti- inflammatory cytokine responses and elucidate a critical role for the GSK3 cell-signaling pathway in modulating the host's inflammatory response to P. gingivalis. The specific aims are designed to characterize how host-microbial interactions are involved in the regulation of the host inflammatory response to P. gingivalis. to elucidate and characterize the involvement of GSK3 in regulating these immune responses, and to evaluate the efficacy of a therapeutic target (GSK3) that could attenuate the disease process in vivo. [unreadable] [unreadable] [unreadable]